111 / 2021-06-30 22:31:26

Improved fenvalerate metabolic activity by point mutations of carboxylesterase 001C from Helicoverpa armigera

Pesticide bioremediation, carboxylesterase, mutation, detoxification

Three major classes of detoxification enzymes, including cytochrome P450s (CYPs), carboxylesterases (CarEs), and glutathione S-transferases (GSTs), are principally responsible for the detoxification of xenobiotic chemical compounds, and their evolution can lead to increased insecticide resistance among insects. Amongst, carboxylesterases (EC3.1.1.1) are a major class of metabolic enzymes belonging to a large superfamily of α/β hydrolase proteins, and play critical roles in the detoxification of toxic chemicals, including organophosphates (OPs) and synthesized pyrethroids (SPs).

Previous work has shown that the CarE 001C from Helicoverpa arimgera is able to metabolize the isomers of two types of insecticides, cypermethrin and fenvalerate. In this study, we have introduced seven mutations into 001C by using site-directed mutagenesis technology, and expressed them in an E. coli system. We purified the recombinant mutant enzymes, determined their kinetic efficiency toward the model substrate α-NA by using spectrophotometric microplate assays, and measured their ability in the detoxification of two types of SPs, cypermethrin and fenvalerate with HPLC method. The results showed that all the mutations in 001C dramatically reduced enzymatic activity toward the generic substrate α-naphthyl acetate, but at the same time increased fenvalerate hydrolase activities in the two cases of H423I and R322L mutation, with the specific activities of 3.5 and 5.1 nM ^-1. min ^-1. mg ^-1 proteins, respectively. These results indicate that these two mutation may have potential to cause fenvalerate resistance in the future.

Apart for the importance in respect of insecticide resistance, the results that the two mutation enhanced the detoxification of fenvalerate also suggested that they have the potential to implicate in pesticide bioremediation in future. It is known that certain detoxification enzyme could be used to decontaminate residues of pesticides in waste waters, contaminated agricultural soils, and so on.